User:Karmella Haynes/Notebook/BioBrick cloning/2015/05/20: Difference between revisions
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|style="background-color: #EEE"|[[Image:owwnotebook_icon.png|128px]]<span style="font-size:22px;">Karmella's BioBrick Cloning</span> | |style="background-color: #EEE"|[[Image:owwnotebook_icon.png|128px]]<span style="font-size:22px;">Karmella's BioBrick Cloning</span> | ||
|style="background-color: #F2F2F2" align="center"| | |style="background-color: #F2F2F2" align="center"|[[File:Report.png|frameless|link={{#sub:{{FULLPAGENAME}}|0|-11}}]][[{{#sub:{{FULLPAGENAME}}|0|-11}}|Main project page]]<br />{{#if:{{#lnpreventry:{{FULLPAGENAME}}}}|[[File:Resultset_previous.png|frameless|link={{#lnpreventry:{{FULLPAGENAME}}}}]][[{{#lnpreventry:{{FULLPAGENAME}}}}{{!}}Previous entry]] }}{{#if:{{#lnnextentry:{{FULLPAGENAME}}}}|[[{{#lnnextentry:{{FULLPAGENAME}}}}{{!}}Next entry]][[File:Resultset_next.png|frameless|link={{#lnnextentry:{{FULLPAGENAME}}}}]]}} | ||
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'''Cas-tone - Phusion PCR for H3''' | '''Cas-tone - Phusion PCR for H3''' | ||
* Phusion PCR-amplify ORFs | |||
# mEmerald-H3-23, H3_1 f1 / H3_40 r1 | |||
# mEmerald-H3-23, H3_1 f1 / H3_60 r1 | |||
# mEmerald-H3-23, H3_1 f1 / H3_80 r1 | |||
# mEmerald-H3-23, H3_1 f1 / H3_116 r1 | |||
# mEmerald-H3-23, H3_1 f1 / H3_136 r1 | |||
{| {{table}} cellspacing="3" <!-- PCR rxn table --> | |||
| bgcolor=#cfcfcf | Reagent | |||
| bgcolor=#cfcfcf | Vol | |||
| bgcolor=#cfcfcf | Mix (x6) | |||
| rowspan=7 | Expected:<br>1. H3-1-40 = 129<br>2. H3-1-60 = 189<br>3. H3-1-80 = 250<br>4. H3-1-116 = 373<br>5. H3-1-136 = 433 | |||
| rowspan=7 | [[Image:KAH052015_gel1.jpg|250px|Hover name]]<br>5 μL/lane; 1% agarose; [http://openwetware.org/wiki/Image:KAH_Fermentas_GeneRuler_1kbplus.jpg Ladder] | |||
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| Plasmid DNA || 0.5 || 3.0 | |||
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| 10 μM H3_1 f1 || 1.0 || 6.0 | |||
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| 10 μM r primer || 1.0 || --- | |||
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| 10 mM dNTPs || 1.0 || 6.0 | |||
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| 5x HF buffer || 10.0 || 60.0 | |||
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| Phusion Pol. || 0.5 || 3.0 | |||
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| dH<sub>2</sub>O || 36.0 || 216.0 | |||
|- | |||
| || 50.0 μL || | |||
|} | |||
Thermal cycler: Bio-Rad - Phusion | |||
* 98°C 3 min. | |||
* 30x[98°C, 10 sec; 70°C 30 sec; 72°C 30 sec] | |||
* 72°C 3 min. | |||
* 4°C, ∞ | |||
Latest revision as of 00:58, 27 September 2017
Karmella's BioBrick Cloning | Main project page Previous entry Next entry | |||||||||||||||||||||||||||||
05/20/15
Cas-tone - Phusion PCR for H3
Thermal cycler: Bio-Rad - Phusion
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